Methicillin resistant Staphylococcus aureus (MRSA) and methicillin sensitive S. aureus (MSSA) are highly significant human health threats that are responsible for a range of diseases. Recently, the CDC and colleagues reported that MRSA are the most significant causes of serious infections and infectious disease deaths in the United States. A major factor involved in the lethality of MRSA and MSSA are secreted toxins, known as super antigens. The lethality associated with these super antigens has been appreciated for decades, as the U.S. military stockpiled large amounts of staphylococcal enterotoxin B (SEB) under its old bioweapons program that was terminated in 1969. The mechanism of action of the toxins is well known - they act extracellularly by binding to the T cell receptor on T cells and stimulating massive release of inflammatory cytokines. We have engineered soluble T cell receptors for high-affinity binding to super antigens to prevent toxin-associated lethality. The objective of this proposal is to build on the success of work completed in Phase I where treatment with the novel therapeutic (soluble, high-affinity T cell receptor, called IMV-01) was able to neutralize SEB secreted from MRSA in an intrapulmonary rabbit model. The hypotheses of the Phase II work are that yields of the therapeutics can be improved, that IMV-01 and two soluble T cell receptor leads (IMV-02 and IMV-03) already engineered against the other clinically important super antigens (SEC & TSST-1) can be co-administered as a cocktail to neutralize these toxins, and prevent lethality due to pneumonia caused by MRSA. The Specific Aims are: 1) To determine the best conditions for optimizing the yield and purity of the V?-TCRs IMV-01, IMV-02 and IMV-03; 2) To determine the effective dose and timing of IMV-01, IMV-02 or IMV-03 individually to neutralize their respective toxins produced by MRSA strains in an intrapulmonary rabbit model in the presence or absence of vancomycin; and 3) To determine the efficacy of using IMV-01, IMV-02 and IMV- 03 as a cocktail to protect rabbits from death due to intrapulmonary challenge with three different MRSA strains secreting either SEB + SEC, SEB + TSST-1 or SEC + TSST-1, in the presence or absence of vancomycin. PUBLIC HEALTH RELEVANCE: Antibiotic-resistant Staphylococcus aureus isolates frequently initiate serious human diseases through the bacterium's colonization of respiratory mucosa and skin, followed by secretion of potent toxins referred to as super antigens. Illnesses include serious post viral pneumonias, allergic skin diseases, and delayed wound healing that affect thousands of individuals each year in the United States. The secreted toxins are directly involved in the effects of the, disease and can cause death. In this application, ImmuVen will develop a therapeutic that neutralizes these toxins, and examine its effectiveness in various models of lethal pneumonia caused by the bacterium.